The contribution of age-related changes in the gut-brain axis to neurological disorders

ABSTRACT Trillions of microbes live symbiotically in the host, specifically in mucosal tissues such as the gut. Recent advances in metagenomics and metabolomics have revealed that the gut microbiota plays a critical role in the regulation of host immunity and metabolism, communicating through bidirectional interactions in the microbiota-gut-brain axis (MGBA). The gut microbiota regulates both gut and systemic immunity and contributes to the neurodevelopment and behaviors of the host. With aging, the composition of the microbiota changes, and emerging studies have linked these shifts in microbial populations to age-related neurological diseases (NDs). Preclinical studies have demonstrated that gut microbiota-targeted therapies can improve behavioral outcomes in the host by modulating microbial, metabolomic, and immunological profiles. In this review, we discuss the pathways of brain-to-gut or gut-to-brain signaling and summarize the role of gut microbiota and microbial metabolites across the lifespan and in disease. We highlight recent studies investigating 1) microbial changes with aging; 2) how aging of the maternal microbiome can affect offspring health; and 3) the contribution of the microbiome to both chronic age-related diseases (e.g., Parkinson’s disease, Alzheimer’s disease and cerebral amyloidosis), and acute brain injury, including ischemic stroke and traumatic brain injury.


Introduction
Over the past decade, research has identified a novel role of the gut microbiome in the bidirectional communication between the gut and the brain, termed the microbiota-gut-brain-axis (MGBA). 1 Disruption in the balance of gut microbial communities (often referred to as "dysbiosis") has implicated various pathways along this axis that contribute to the progression of neurodegenerative diseases, such as Alzheimer's disease (AD) and Parkinson's disease (PD). 1 The MGBA also contributes to outcomes after acute neurological injury, such as stroke and traumatic brain injury (TBI). 1 Many of these NDs are diseases that increase in prevalence with aging.Emerging studies have shown that the process of aging directs and changes the composition of the microbiome, 2 which leads to chronic systemic inflammation, or "inflammaging". 3Inflammaging is characterized by an increased level of circulating proinflammatory cytokines and breakdown of the barrier integrity of host tissues including in the brain (e.g., blood-brain barrier) and gut (e.g., intestinal epithelium), leading to antigen translocation into the host and heightened systemic inflammation. 2,4nflammaging may be caused by a variety of processes that accompany aging, such as oxidative stress or cellular senescence, many of which are mediated by recently identified pathways in the MGBA. 5 In experimental studies, specific microbially-derived metabolites are altered both with aging and in agerelated diseases, which has now been confirmed in clinical populations as well. 6Our understanding of the mechanisms through which these metabolites change host homeostasis is emerging, but detailed mechanistic studies are required if we hope to harness the potential of the microbiome to enhance health.effect on all physiological and pathological processes occurring in the host. 1,7Microbial communities vary across distinct body sites and organs, but those residing in the gastrointestinal (GI) tract have, to date, attracted the greatest attention in biomedical research, including gerontology and neurological research.
Multiple large-scale studies, such as the NIHfunded Human Microbiome Project (HMP), have provided extensive information regarding the genetic sequences of most microorganisms residing in the human gut.These datasets revealed that the phyla Bacteroidetes and Firmicutes account for 90% of the gut microbiota, but many others such as Proteobacteria, Actinobacteria, Fusobacteria, Spirochaetes, Verrucomicrobia and Lentisphaerae are also present.The identification of a core microbiome, despite being critical for our understanding of the microbial contribution to health and disease, is challenging due the huge variability in gut microbiome configurations across individuals and within the same subject across the lifespan. 8More recently, multi-omics approaches such as metagenomics and metabolomic sequencing has dramatically expanded our knowledge of the functional role of gut microorganisms in host health and pathological processes. 9The interplay between the gut microbiota and the host is regulated by a complex network of metabolic, immune, and neuroendocrine interactions.When physiological changes within the gut microbial community evolve into a detrimental state, or so-called "dysbiosis", 10 significant alterations in the pool of the metabolites produced and released by these microorganisms occur, with important repercussions for host physiology.
A growing body of evidence has implicated the gut microbiome in the progressive accumulation of molecular and cellular alterations observed with senescence.These changes ultimately increase the susceptibility to chronic diseases in older populations.Aging-associated processes include the progressive accumulation of senescent cells (SCs), which are identified by definitive cell cycle arrest, abnormal mitochondrial reactive oxygen species (ROS) production, metabolic shifts, and the production of senescence-associated secretory phenotype factors (as reviewed in ref 11 ).Release of these factors triggers proinflammatory responses from different immune cells that participate in the physiological changes seen with aging.These detrimental changes are potentially reversible, as shown by recent studies in mice that target these SCs, leading to reductions in systemic inflammation, TNFα/NF-κB signaling, and senescence-associated signatures in aged mice. 12Intriguingly, emerging evidence suggests that the gut microbiome may play an important role in modulating the effects of SC.A very recent study using germ-free (GF) mice (raised in total absence of microbial colonization) showed that the aging microbiome was responsible for accumulation of senescent markers in ileal B cells, which in turn further altered gut microbiome composition. 13

The gut-brain axis: a bidirectional communication
Communication between the brain and microbiota is bidirectional and can occur through multiple pathways.These include neural connections between the brain and the gut through the vagus nerve, hormonal and immune pathways, and metabolite signaling. 14There is a high degree of intercommunication between the gut and the peripheral nervous system (PNS) which participate in the immunological and hormonal responses to gut bacterial biochemical processes.Gut microbial signals can be "sensed" via vagal and spinal neurons, integrated in the brainstem and hypothalamus, and ultimately influence efferent signals to peripheral organs. 14Several recent studies manipulating the gut microbiota composition have illustrated the importance of the interaction between gut microbes and the PNS (Figure 1), via efferent/afferent pathways, in regulating host physiology, as discussed below.

Neuroanatomic pathways: vagal mechanisms/ vagus nerve
The two-way neuroanatomic communication between the gut and brain occurs through afferent or efferent signaling along two main directional pathways: (1) the autonomic nervous system (ANS) including the vagus nerve (VN) and the enteric nervous system (ENS).The VN is the tenth cranial nerve and one of the main components of the parasympathetic nervous system, which forms the ANS together with the sympathetic nervous system.The ANS has a primary role in regulating multiple physiological processes, including heart rate, immune response, and digestion. 15Signals from the gut are conveyed to the central nervous system (CNS) through the ANS in a bottom-up manner, and responses from the CNS are then sent to the gut by the ANS in a topdown manner. 16In this context, the VN represents the most direct connection between the gut and the brain, participating in both bottom-up and topdown signaling via both afferent (sensory) and efferent (motor) nerves. 17Vagal terminals reach the gut in the mucosal layer, the smooth muscle layer, and synapse with enteroendocrine cells (EECs), without direct contact with the gut microbiota in the lumen.A recent study by Bohórquez and colleagues 18 showed the presence of a specialized group of EECs, defined as neuropods, which provide a direct connection between the gut lumen and brain stem by synapsing with the VN through glutamatergic transmission.The neuropod-vagal terminal circuit is activated in response to sugar, thereby transducing fast sensory input from the gut lumen.Additionally, vagal fibers, which express receptors for multiple metabolites produced by the microbiome, can sense changes in microbial populations. 19Similarly, EECs express receptors for microbial metabolites such as short-chain fatty acids (SCFAs), indoles, bile acids, and lipopolysaccharide (LPS). 20Both human and animal studies have highlighted the crucial role of the VN in regulating brain activity.Either partial or total vagotomy in rodents led to changes in brain circuits and behavioral functions implicated in various neuropsychiatric disorders, 21 such as anxiety, fear-related phenotypes, 22 learning and memory, 23 locomotion 24 and sensorimotor gating. 25Similarly, direct stimulation of the VN modulates stress-induced depressive phenotypes via regulation of serotonergic circuitry in the hippocampus, 26 in anxiety and post-traumatic stress disorders 27,28 and the reward system involved in affective disorders through effects on dopaminergic circuitry in the substantia nigra. 29ecent studies have identified specific microorganisms residing in the gut that can modulate brain function via vagal fibers.For example, administration of Lactobacillus rhamnosus was effective in decreasing both depressive-and anxiety-like phenotypes in mice, an effect that was mediated by an increased firing rate of VN terminals. 11Similarly, supplementation with Campylobacter jejuni increased VN c-Fos expression in the vagal ganglia, leading to activation of neurons in the solitary nucleus of the brainstem. 30Additionally, another study showed that C. jejuni treatment in mice promoted anxiety-related behavior. 31The interactions between the vagus nerve, the gut epithelium and the ENS are summarized in Figure 2.

Neuroanatomic pathways: the enteric nervous system
The ENS, a part of the PNS, is at a critical intersection between the host and the gut microbiome.Anatomically, the ENS is organized as a web of motor, sensor, and interneurons that are embedded in the inner and outer layers of the muscularis ] externa and in the submucosa of the digestive sys- tem.By integrating peripheral sensory information with input from the ANS, the ENS controls the muscular and secretory functions of the GI tract, including peristalsis and the production and release of enzymes and hormones, such as gastrin and secretin. 28Sensory neurons in the ENS form synapses with both enteric motor neurons and vagal fibers, and express receptors for multiple microbial metabolites and components, including SCFAs 32 and LPS through toll-like receptor 4 (TLR4). 33Studies in GF mice have shed light on the influence of the microbiome in controlling the electrophysiology of ENS neurons.Maturation of the ENS begins during postnatal development as microbial strains colonize the infant gut through the activation of pattern recognition receptors, such as TLRs, on ENS terminals that bind to microbial products, including LPS. 34 Microbial reconstitution of GF mice induced upregulation of the expression of 5-hydroxytryptamine (5-HT) and its receptors in enteric neurons. 34,35Antibioticmediated depletion of gut bacteria in mice altered both the morpho-functional structure and the neurochemistry of the ENS, including the loss of neurons in the myenteric plexus, increased TLR2 expression in neuromuscular and mucosal layers in the ileum, and a reduction in glial cells. 36ifferent microorganisms may exert different effects on ENS neuronal activity through distinct mechanisms. 37Recent studies suggest that changes in the ENS might be a driving factor in determining dysbiosis of the gut microbiome through the regulation of intestinal transit, gut barrier permeability, and luminal pH. 38

Systemic and mucosal immune regulation: immunological pathways
Studies in both animal models and humans have shown that the gut microbiome is essential for the regulation of the host immune system.Microbial colonization of the host's mucosa during the early postnatal period profoundly shapes the development and maturation of the host immune system. 39Beyond infancy, the gut microbiome is intricately involved in maintaining immune homeostasis through complex interactions with the mucosal immune system.
Immune cells also play important roles in the gut 40 including: (1) tolerance toward a multitude of microorganisms in the normal, healthy gut ecology; (2) surveillance of potential pathogenic strains; and (3) inhibition of commensal overgrowth and prevention of translocation of bacteria from the intestinal lumen into the host, a process that requires the integrity of the intestinal mucosal barrier. 41rosstalk between the gut microbiota and host immune system regulates the production and release of neurotransmitters and neuropeptides, cytokines, and other signaling mediators that influence brain function by multiple mechanisms, including actions on vagal and spinal afferent fibers. 42In this context, one of the mechanisms of regulation of the MGBA involves the maturation and function of microglia, the resident immune cells of the brain. 42,43This process starts during fetal and early postnatal development and is mediated by the composition of the maternal gut microbiome. 44uring prenatal development, maternal gut microbial communities contribute to fetal microglia programming, which in turn, directly affect the formation of cortical cytoarchitecture and neural circuits. 45In GF mice, substantial alterations in gene expression occur in fetal microglial at mid and late gestation.Microglia programming continued postnatally (P20) leading to increased microglial density (Iba1 + cells) in the somatosensory neocortex of GF females compared to SPF females, which was not seen in males. 44The impaired microglia maturation seen in GF mice was rescued by early-life colonization with a complex microbiome. 46Migration of CD4 T cells into the brain around the time of birth is also critical for microglial maturation, and is potentially regulated by the gut microbiome. 47Therefore, further studies are needed to investigate how alterations in the gut microbiome composition and immune cell populations in early development could lead to increased predisposition for neurological disorders later in life.
Microglia not only modulate inflammatory processes in the brain, but are also involved in synaptic plasticity and remodeling, maturation of the CNS, and debris and aggregate clearance. 48Microglia activation and function can be mediated by several factors produced from host-bacteria interactions, 49 including cytokines, tryptophan metabolites, bacterial-derived cell components (e.g., peptidoglycans and LPS) 50 and bacterial-derived metabolites (SCFAs). 51These gut-derived signals can reach the brain through the bloodstream, through the VN 52 and potentially through the newly discovered meningeal lymphatic system through actions on γδ T cells. 53Several studies have proposed epigenetic regulation and chromatin remodeling as a potential mechanism for microbiota-dependent modulation of microglia function. 43,54A recent investigation in the context of neurodegenerative processes 55 identified SCFAs, specifically acetate, as a crucial regulator of microglial metabolic function through histone methylation modification on genes related to microglial proliferation, morphology, activation, and metabolism.Epigenetic regulation of immune function via bacterial metabolites has also been observed in the gut immune system.For example, SCFAs, particularly propionate, decreased IL-17-producing γδ T cells in humans (e.g., peripheral blood mononuclear cells) and in mice (e.g., intestinal lamina propria) in a histone deacetylase-dependent manner. 56Further research is warranted to unravel the precise mechanisms by which the gut microbiome modulates microglia and, as such, brain function.However, these studies have provided mechanistic insights that might lead to the identification of new microbiome-based therapeutic strategies that target microglial function in brain disorders.

Neuroendocrine-hypothalamic-pituitaryadrenal axis pathway
The neuroendocrine system regulates many processes in the human body and plays a critical role in organ development and function.Among the primary neuroendocrine pathways, the hypothalamic -pituitary -adrenal (HPA) axis is thought to be tightly connected to the gut microbiome in the context of the MGBA.The HPA includes the hypothalamus, the pituitary gland, and adrenal glands. 57In the context of the stress response, the hypothalamus receives stimuli to produce and release corticotrophin-releasing hormone (CRH), which induces the secretion of adrenocorticotropic hormone (ACTH) from the pituitary gland.ACTH stimulates the adrenal cortex to release glucocorticoids, mineralocorticoids, and catecholamines, which then modulate other downstream processes to produce appropriate responses to the stressor. 58idirectional communication between the neuroendocrine-HPA axis system and the gut microbiota involves multiple other components of the MGBA, including the immune system, gut hormones and microbially-derived products, as well as receptors expressed on both the intestinal and the blood-brain barriers as reviewed in ref. 58 Several studies have suggested that microbiallyderived compounds, such as precursors of neurotransmitters and gut hormones, and SCFAs, can regulate the neuroendocrine system. 58,59Similarly, the microbiome may participate in HPA axis modulation through the immune system. 60yperactivation of the HPA axis is associated with disorders affecting both the neuroendocrine system and the gut microbiome, such as irritable bowel syndrome (IBS) and depression.One intriguing hypothesis 61 suggests that HPA axis dysregulation might lead to gut dysbiosis and alterations in the integrity of the intestinal barrier, which in turn could promote chronic low-grade inflammation that is seen in both IBS 62,63 and depression. 64,65he microbiome is involved in the bidirectional communications between the neuroendocrine system, including the HPA axis, and the immune response 66,67 as reviewed in ref. 68 The integrity of the intestinal barrier can be altered by neuroendocrine mediators released in the context of stress response, thus facilitating the release of microbially-derived molecules, which subsequently activate immune pathways.69 For instance, levels of bacterial LPS increase the production of cytokine colony-stimulating factor 1 in muscularis macrophages in the myenteric plexus, which leads to regulation of gut motility, such as increased colonic transit time through the ENS.70 Similarly, alterations in the gut microbiome and its metabolites can induce activation of the HPA axis and associated immune responses.71 Studies in GF mice showed that the absence of gut microorganisms is linked to neuroendocrine and behavioral alterations.72,73 GF mice showed exaggerated HPA axis activation in response to stress, with increased levels of circulating corticosterone, and independent of cytokinemediated pathways, as shown by unaltered IL-1β and IL-6 levels in the plasma.Intriguingly, colonization with specific strains, such as B. infantis or E. coli, modulated the HPA axis by either increasing or decreasing its activity, respectively.73

Microbially-derived neuroactive metabolites
The MGBA is regulated by a large number of different neurotransmitters, neuropeptides, and microbially derived products. 74While neurotransmitters have multiple effects on gut ecology, the microbiome itself produces and releases neurotransmitters.Studies showing a microbial origin for dopamine, norepinephrine, gamma-aminobutyric acid (GABA), and serotonin, among others, point toward a potential role of microbiota-produced neurotransmitters in influencing brain function, as discussed below. 75dditionally, neuropeptide synthesis is influenced by hormones and amino acid availability, which is controlled by the microbiome. 74However, neuropeptides can also modulate the composition and function of the gut microbiota, reflecting the complex bidirectional crosstalk that integrates these systems.
Bacteria have unique structural components known as microorganism-associated molecular patterns (MAMPs), such as LPS.MAMPs play an important role in host development and immune function. 76Transformation of host-derived components involves the production of secondary bile acids and steroid hormones, which have neuroactive properties. 77The gut microbiota also plays an important role in the transformation of dietary substrates.Many microbially-derived metabolites of amino acids, carbohydrates, and other plantderived molecules exert pleiotropic effects on the MGBA and ultimately, on brain function and behavior.The microbial metabolism of tryptophan, tyrosine, and phenylalanine influences the production of neurotransmitters such as serotonin, noradrenaline, and dopamine. 78Tryptophan is particularly relevant to the brain, including tryptophan derivatives, indoles and kynurenine, which can modulate glutamate signaling and have been implicated in anxiety-like behavior and cognitive dysfunction. 79,80The microbiome is also implicated in the production of the major inhibitory neurotransmitter GABA, and alterations in glutamate/GABA circuits in the brain have been associated with the development of autism spectrum disorders, schizophrenia, major depressive disorder, and other neuropsychiatric disorders. 11For instance, a wide analysis of GABA production levels from several commercially available probiotics highlighted the strain-specific ability of Levilactobacillus brevis and Lactiplantibacillus plantarum in secreting GABA both in vitro and in vivo. 81Additionally, expression of GABA receptors can be modulated by specific bacterial strains such as Lactobacillus rhamnosus in mice, with data indicating a decrease in GABA B1b mRNA in the hippocampus and amygdala, and concomitant decrease in GABA Aα2 mRNA expression in the prefrontal cortex.These changes were associated with reduced anxiety-and depression-like behaviors. 11omplex plant polysaccharides, or dietary fibers, are fermented by the gut microbiome to produce SCFAs, a class of compounds that has recently gained a great deal of attention because of their ability to influence multiple processes in the host, including behavior. 82,83SCFAs, such as butyrate, propionate, and acetate, are used as an energy source for colonic epithelial cells and can enter the systemic circulation and modulate the immune system through the regulation of gene expression. 84CFAs can also cross the BBB via monocarboxylate transporter-expressed endothelial cells, where they can directly act on both neurons and microglia (as extensively reviewed in ref 84 )in the neurological and neuropsychiatric disorders investigated.Recent findings on the role of neuroactive metabolites and their effects on the MGBA 21,[85][86][87][88][89][90][91][92][93] are summarized in Table 1.In the following section, we will focus on the link between gut dysbiosis and neurological disorders in the context of the MGBA.

Microbiome changes with aging
Microbial colonization of the GI tract begins at birth, with mother-to-infant vertical transfer of skin and vaginal microbial strains, which are then replaced by species typically seen in the adult gut.The first 1,000 days of life are characterized by a relatively low bacterial diversity, with genus Bifidobacterium representing up to 50% of the infants' gut microbial community, followed by an intense remodeling of the foundational gut microbiota. 94Fluctuations in the infant microbiome configuration follow the transition from a breast milk-based diet to an adult-like diet at the time of introduction of solid food and are also influenced by the surrounding environment. 95iven the instability of the infant gut microbiota, exposure to detrimental environmental factors can disrupt this highly orchestrated microbial succession, leading to gut dysbiosis that persists well beyond the early developmental period and serves as a risk factor for disease later in life. 96This first period of microbial colonization is characterized by rapid changes determined by either environmental factors or intrinsic ecological drifts which continue throughout adolescence.In contrast, relative stability in gut microbial communities is reached during adulthood.Even though environmental factors, such as antibiotic treatment and changes in diet, can alter gut microbiome composition, key species are thought to regulate the integrity and stability of the ecosystem in adult individuals.
Aging is a complex, time-dependent decline of the physiological, immunological, metabolic, and genomic functions in the host.Many studies 97 have attempted to describe the molecular and cellular hallmarks of aging, which include cellular senescence, telomere dysfunction and damage, alterations in protein synthesis and epigenetic regulation, mitochondrial and nutrient-sensing dysfunction, and depletion of stem cell reserves.More recently, chronic inflammation and gut dysbiosis have emerged as additional factors associated with aging. 98Immune aging is characterized by "immunosenescence," a progressive decline in the ability of both innate and acquired immunity to induce an effective response to both infection and vaccination. 99hanges in the gut microbiome are implicated in both age-related pathologies and potentially act as mediators of the aging process. 100 major change seen in the gastrointestinal tract with aging is a decrease in the barrier integrity of the intestinal epithelium. 2 The function of the intestinal barrier can be modulated by commensal resident microbiota.Akkermansia, for example, given orally to mice has been shown to alleviate senescence-related phenotypes in the intestines of aged mice. 101In young mice, higher amounts of Parabacteroides and Akkermansia were found, whereas these two bacterial taxa decreased in the gut of aged mice. 101kkermansia induces mucus production in the gut, which may help to restore and maintain barrier integrity. 102The effects of aging on the microbiome also contribute to changes in pathways controlled by the gut microbiome, including those involved in the biosynthesis of GABA and SCFA production, which are less enriched in aged mice than in young mice. 101In humans, Bifidobacterium are found at higher levels in infants, while Lachnospiracae levels are higher in adults. 103However, some consistencies remain throughout a healthy lifespan, evident through data that show how certain bacterial species, such as Enterobacteriaceae, are found at similar levels in both infants and the elderly. 103Some researchers have suggested that changes in the microbiota that occur with aging may be better assessed using biological age, rather than chronological age, as the contributing factor.Although methods for determining biological age can produce varying results, researchers have found that the Frailty Index (FI34), a method to calculate biological age, is better correlated with changes in microbiota in humans than chronological age. 104ne method of identifying and characterizing biological age may be through characterization of the gut microbiome composition.As implicated in many studies 6,101,103,105,106 (Table 2), the gut microbiome composition is significantly altered with increasing age, even in the absence of an ND, which further exacerbates gut dysbiosis.

The role of an aging maternal gut microbiome in offspring neurodevelopment
The maternal exposome 107 in particular, plays a crucial role in early life development. 108etrimental alterations in the maternal exposome can trigger fetal programming events that predispose offspring to chronic health conditions, including brain disorders, later in life ('Developmental Origins of Health and Disease' 109 ) (Figure 3).][117][118][119] This may involve epigenetic reprogramming in either the oocyte 120 or the fetus, or direct effects of inherited dysbiosis.The precise mechanisms by which AMA affects brain development are unclear. 114Over the past three decades in the United States, there has been a steady increase in the birth rate for women aged 35-39 years from 45.9 per 1000 women in 2010, to 52.7 in 2019. 121imilarly, the birth rate for women aged 40-44 years rose by 5% from 2020 to 2021. 122Although a multitude of factors contribute to the increased risk of complications seen in older mothers and their offspring, recent studies have implicated changes in the maternal microbiome that may contribute to these poor outcomes. 123,124espite the established association between aging, gut dysbiosis, and increased inflammation, few studies have focused on the effects of maternal age-related gut dysbiosis on fetal development and brain health outcomes.A recent study in humans showed that both the vaginal and the gut microbiomes of women displayed significant differences in microbial composition based on age and pregnancy status. 125Given that aging is characterized by gut dysbiosis, an altered intestinal metabolome, increased barrier permeability, and chronic, lowgrade inflammation, it is possible that advanced maternal age could alter both the maternal gut microbiome and gut mucosal and systemic immune system, similar to what is observed in maternal obesity.This can then disrupt physiological adaptations to pregnancy and impair placental function, leading to increased brain and systemic inflammation in the fetus and alterations in neurodevelopment (Figure 4).These effects might be mediated by gut dysbiosis-mediated alterations in the abundance of microbiomederived metabolites, such as SCFAs, [126][127][128] which can (1) actively modulate maternal immune cells, leading to enhanced systemic and placental inflammation 129 and (2) cross the placenta and directly influence epigenetic programming of fetal brain cells, neuroinflammation through microglial activation, and neural circuit formation.After birth, maternally inherited dysbiosis can sustain systemic and neuroinflammatory events in offspring, leading to detrimental effects on brain function and behavior.It is unknown how long these detrimental changes last in the offspring, or if these early developmental events can alter the risk for neurodegenerative diseases later in life.Longitudinal studies are required to address this question.

Parkinson's disease (PD)
PD is a neurodegenerative disease characterized by a loss of dopaminergic neurons in the substantia nigra pars compacta and the deposition of insoluble alpha-synuclein polymers in neurons, forming Lewy bodies. 130pproximately 80% of PD patients suffer from GI dysfunction. 131PD patients commonly suffer from symptoms such as constipation, which precede the clinical diagnosis of PD and its other hallmark symptoms such as bradykinesia and dementia, 131 indicating that gut dysfunction may play a role in the pathogenesis of PD.Recent studies have attempted to identify specific microbiota changes that may lead to PD.An MPTP-induced mouse model of PD showed very distinct changes in the gut microbiome, including a significant decrease in the levels of Prevotella and Faecalibacterium and an increase in Ralstonia bacteria, compared to control mice. 132Additionally, Enterobacteriaceae is increased in both humans and rodent models of PD. 133 The specific mechanisms behind these microbiota changes and their contribution to PD are yet to be understood.However, certain bacterial species present in the gut or fecal matter of patients with PD have been welldescribed (Table 3).Citrobacter rodentium, which is enriched in patients with PD, has also been shown to aggravate motor symptoms in mouse models.Increases in Proteus mirabilis are also linked to PD symptoms and have been shown to promote motor deficits in mouse models of PD. 139 Researchers have linked the changes in the expression of these specific bacterial genes to mechanisms that regulate lipid biosynthesis and secretory pathways, including dopamine (DA) regulation and production, as many PD symptoms can be traced back to a decrease in DA levels. 140Studies have shown that some of the bacterial enzymes residing in the gut produce DA, 141 further solidifying the link between the roles of microbial-derived metabolites in the progression of PD.This direct correlation between bacterial metabolites and the onset of PD symptoms implies there a role of the MGBA in the pathogenesis of PD (Table 3) 134-138; however, the specific metabolites involved remain to be studied in-depth.

Alzheimer's disease (AD)
One of the most common diseases associated with aging is AD; increasing age is the greatest risk factor for late-onset AD. 142 Although some treatments targeting amyloid clearance in AD patients have emerged, the availability of therapeutics targeting the prevention of amyloid development is limited. 143Interestingly, recent studies have demonstrated that amyloid-beta (Aβ) plaque deposition is linked to the composition of the gut microbiota.Studies by our research group have shown that in a Tg2576 transgenic mouse model of AD, gut inflammation and dysbiosis precede the accumulation of amyloid plaques in the brain, 144 indicating that gut dysbiosis may play a role in the development of amyloid pathology, although these (age:69.3± 7.5 years). 145Liu et al. also observed a decrease in Firmicutes and an increase in Proteobacteria in the elderly patients. 146In addition, Cattaneo et al. showed that the abundance of proinflammatory bacteria such as Escherichia/Shigella is increased in patients, whereas that of an antiinflammatory bacterium (e.g., E. rectale) is increased. 147These findings indicate that gut microbiota may be associated with AD pathophysiology. 148ther studies (Table 4) 145,146,[149][150][151][152] show that among different species and disease models of AD, many of the mechanisms or changes governing microbiota-driven alterations in AD development are similar.Some of these changes, however, appear to be species-dependent, that is, they are observed differently in humans and animal models.
To understand the role of gut microbiota in AD, microbiota-targeted interventions, including fecal microbiota transplantation (FMT), have recently been employed in animal models of AD.Sun et al. performed FMT from naïve WT mice (6 months old) into agematched APPswe/PSEN1dE9 transgenic mice.These transgenic mice exhibit occasional Aβ deposits by six months and abundant plaques by nine months. 153Cognitive impairment is seen at 12-13 months in this mouse model. 153,154They found that FMT improves cognitive function and synaptic plasticity, and decreases levels of Aβ40, Aβ42, and p-Tau231 in the brain of recipient mice. 155The beneficial effects of FMT seen in the recipient mice were associated with higher levels of fecal SCFAs, such as butyrate.More recently, Kim et al. transplanted the fecal microbiome of 5×FAD mice into WT mice. 156Compared with many other models, 5×FAD mice show more rapid Aβ deposits in the brain (<3 months) and cognitive impairment (<6 months). 157,158They showed that reconstitution of the 5×FAD microbiome reduced spatial learning and memory in recipient WT mice, compared with recipient mice treated with the biome from WT mice.In addition, recipient mice with the 5×FAD microbiome showed decreased neurogenesis, increased neuroinflammation including microglial activation, and elevated proinflammatory cytokines (e.g., TNF-α and IL-1β) in the brain.Interestingly, the recipient mice had increased levels of both proinflammatory cytokines (e.g., TNF-α, IL-1β, and IL-6) and anti-inflammatory cytokines (e.g., IL-10) in the colon, whereas only IL-1β, but not the other tested cytokines, was increased in the plasma.This indicates that the reconstitution of the gut microbiota with healthy microbiota can ameliorate memory dysfunction by regulating inflammation in AD mice through the MGBA.It has also been reported that transferring healthy microbiota into ADLP APT mice, a mouse AD model with both amyloid and neurofibrillary tangle pathology, significantly reduces the formation of amyloid plaques and tangles, resulting in cognitive improvement. 151Taken together, these findings indicate that the restoration of a healthy biome can delay the symptoms and progression of AD in animal models.Thus, future investigations of the role of gut microbiota as new therapeutic targets for AD are warranted.

Cerebral amyloidosis and cerebral amyloid angiopathy (CAA)
Amyloidosis, one of the most significant pathologies found in AD, is also affected by gut dysbiosis in related neurodegenerative disorders, including CAA.CAA is a small vessel disease characterized by amyloid deposition in the basement membrane of the brain vasculature. 159Aging is a major risk factor for CAA; CAA leads to progressive cognitive impairment in elderly patients, and also contributes to ischemic small vessel disease and intracerebral hemorrhage. 160n a mouse model of APP and PS1 mutations (APP/ PS1 mice), Chen et al. showed that the microbiota composition between APP/PS1 and WT mice diverged significantly at 1-3 months of age, prior to the onset of cognitive symptoms, amyloid deposition, and neuroinflammation (e.g., microglial activation) in the brain. 161This study, consistent with many others, demonstrated that higher levels of Enterobacteriaceae, as well as Verrucomicrobia were present in the gut of mice that developed amyloid plaques when compared to control mice. 161,162There are very limited studies demonstrating the regulatory role of gut microbiota in CAA or cerebral amyloidosis.Therefore, the investigation of how vascular Aβ versus parenchymal Aβ affects the gut microbiota and gut dysbiosis-associated cognitive impairment in the context of CAA or cerebral amyloidosis will be an important future direction.

Stroke
Stroke is a leading cause of mortality and morbidity in elderly patients.Options for acute treatment such as recombinant tissue plasminogen activator and endovascular thrombectomy are available, 163 however post-stroke treatment is critical as chronic disability and other long-term health consequences of stroke persist for decades. 164The majority of strokes are caused by occlusion of an artery, either by an embolus or an in-situ thrombosis, leading to an area of brain ischemia. 1657][168] In a study comparing young, stroke mice to uninjured aged controls, we found that the gut microbiome of stroke mice is altered and resembles the microbiome composition of uninjured aged mice. 169It was previously reported that post-stroke translocation of gut microbes into the lung leads to sepsis in mice. 1707][168] Using mouse models of ischemic stroke, such as middle cerebral artery occlusion (MCAO), Singh et al. found that stroke can cause gut dysbiosis, as assessed by reduced bacterial diversity and Bacteroidetes overgrowth, which were associated with impaired gut integrity and motility. 166They subsequently transplanted post-stroke microbiome into GF mice.Interestingly, recipient GF mice had larger infarct volumes and worse behavioral deficits, along with increased pro-inflammatory T cells in both the intestines and the ischemic brain.Furthermore, Benakis et al. revealed that gut microbiota can regulate T cell trafficking from the gut into the leptomeninges after stroke and that specific types of T cells, such as regulatory T (T reg ) cells and IL-17 + γδ T cells, are critical in regulating neuroprotection by modulating gut-to-brain signaling following stroke. 168s the elderly are more prone to stroke than younger populations, our research group has focused on the regulatory role of MGBA and the underlying mechanisms of stroke in aged mice.We first examined whether aged mice were more susceptible to stroke-induced gut permeability and bacterial translocation than young mice.Aged mice had increased gut permeability after stroke and higher mortality compared to young mice.When we orally gavaged young and aged mice with GFP-tagged E. coli, aged mice exhibited increased bacterial translocation into peripheral tissues, such as the mesenteric lymph nodes, compared with young mice, 171 indicating the direct effect of aging on gut dysbiosis and bacterial translocation after stroke.
To profile the effect of aging on the composition of the gut microbiota, we performed 16S rRNA-seq on fecal samples from young and aged mice.We found that the composition of the gut microbiota is distinct between young and aged mice; the Firmicutes: Bacteroidetes ratio was higher in the aged biome compared to the young biome, indicating ageinduced gut dysbiosis.Next, we transplanted the aged microbiome and young microbiome into young and aged mice, respectively, using FMT prior to stroke (transient MCAO).Interestingly, young recipient mice with aged microbiome showed increased post-stroke mortality and functional deficits.Conversely, aged recipient mice transplanted with young microbiome showed better post-stroke outcomes. 169Although we found that preconditioning of the aged gut using a young microbiome prior to stroke can contribute to post-stroke recovery, stroke is not predictable.Therefore, in a separate study, we transplanted young microbiome into aged mice several days after stroke (as a treatment) to determine whether post-stroke FMT can improve post-stroke recovery. 126We found that post-stroke reconstitution of young microbiome significantly improved functional outcomes (e.g., increased spontaneous locomotor activity and cognitive functions, and reduced depressive-like phenotype) along with decreased inflammation in both the brain and gut.Post-stroke, young FMT increased T reg cells in the small intestine and enhanced protective mucus production in the large intestine.Moreover, aged stroke mice with young FMT had higher T reg cells and lower IL-17 + γδ T cells in the ischemic hemisphere than aged stroke mice with aged FMT.Using metabolomic analysis, we revealed that the young microbiome contains higher levels of SCFAs, such as acetate, butyrate, and propionate.Based on our metagenomic data, we selected four SCFA producers (Bifidobacterium longum, Clostridium symbiosum, Faecalibacterium prausnitzii and Lactobacillus fermentum) and orally gavaged aged mice with these, and the pre-biotic inulin, after stroke.Interestingly, post-stroke bacteriotherapy using SCFA-producers and inulin increased SCFA levels in the gut, plasma, and brain, and synergistically improved post-stroke recovery by reducing IL-17 production in γδ T cells in the brains of aged stroke mice.In a follow-up study, we found that the aged microbiome alone is sufficient to produce cognitive decline in young GF mice compared to the young microbiome.In conclusion, our findings suggest that aging should also be considered as a detrimental factor regulating the MGBA in stroke.Several other studies 166,[172][173][174][175] (Table 5) suggest that the gut microbiome is an essential regulator of poststroke recovery, and the identification of specific bacterial populations after stroke may uncover various mechanisms through which the microbiome influences inflammation post-stroke.

Traumatic brain injury
Approximately 2 million people sustain a head injury annually in the United States. 176Along with many other symptoms of traumatic brain injury (TBI), intestinal dysfunction has emerged as a chronic consequence of head injury.Studies in rat models of TBI show a loss of alpha diversity and alterations in bacterial taxa that reside in the gut. 177These changes have also been observed in human fecal samples collected from athletes or trauma patients who sustain head injuries or concussions.One such study involving football players who had concussions demonstrated changes in specific bacterial species, such as Agathobacter and Ruminococcaceae, when compared to healthy, uninjured control  subjects. 178Similar results were found in other studies [177][178][179][180] and are reviewed in Table 6.The effect of TBI on the gut microbiome is immediate.The intestinal microbiota becomes disrupted within hours of injury and can lead to chronic inflammatory processes. 181The acuity of post-TBI gut microbiome changes provides evidence that microbiome-targeted therapies could be beneficial for TBI and related head injuries.Therapies targeting microbial alterations that occur with TBI could alleviate the chronic effects of the injury by limiting downstream consequences at the start of injury progression.A summary of the potential role of the gut microbiota in NDs is provided in Figure 5.

Conclusions and future directions
Considerable progress has been made in our understanding of the role of gut microbiota and their metabolites in health and disease.In this review, we have summarized key findings demonstrating the regulatory role of microbiota in neurodevelopment, neuroinflammation, and behaviors of the host, specifically in aging and age-related NDs.
Although some changes in the gut microbial composition vary depending on the context, and substantial limitations (e.g., discrepancy between preclinical animal studies, differences in the gut microbiota composition between animals and humans, and variations in microbiome sequencing and bioinformatic pipelines) still remain, it is accepted that gut microbiota and metabolites are targetable, suggesting that there are novel therapeutic options for NDs through manipulation of the MGBA 155,[182][183][184][185][186][187] (Table 7).Of note, the microbiome has the potential to transform preventative care and reduce medical costs by enabling individual therapies in the field of precision medicine. 188,189Future studies will highlight bacterial strains, metabolites, and immune factors that might help identify new cellular and molecular targets for diagnostic tools and microbiometargeting therapeutic and preventative approaches.

Figure 2 .
Figure 2. Vagus nerve interactions with the gut epithelium and the ENS.Created in Biorender.com.

Figure 3 .
Figure 3. Environmental factors causing detrimental alterations in the maternal exposome.Toxin and pollutant exposure, infection during pregnancy, diet and metabolic status, smoking, psychosocial stressors such as low socioeconomic status, major life events, and pregnancy-related stressors, can determine broad changes in the maternal environment, thereby jeopardizing pregnancy outcomes and fetal developmental programming.Created in Biorender.com.

Figure 4 .
Figure 4. Proposed mechanism for AMA-related fetal programming and increased risk for brain disorders in offspring.AMA-associated gut dysbiosis and increased inflammation may drive abnormal immune activation in both the placenta and the fetal brain, specifically in microglial cells.Therefore, increased brain inflammation could alter neurodevelopment through multiple mechanisms, including epigenetic modifications in neuronal and glial cells.At birth, vertical transmission of a dysbiotic gut microbiome sustains this systemic neuroinflammation in the neonate, jeopardizing postnatal neurodevelopment and adult health.Created in Biorender.com.

Figure 5 .
Figure 5. Major symptoms and features of NDs are accompanied by multiple alterations in specific microbial species, changes which can be consistent or contradictory the human microbiome and mouse models.Created in Biorender.com.

Table 1 .
Neuroactive metabolites and their effects.
101adult offspring.Additional epigenetic programming during early development can also be passed on to subsequent generations.

Table 7 .
Microbiome-targeted treatments for NDs.169middle-aged and older adults with MCI randomized to either probiotic (Lactobacillus rhamnosus) or placebo treatments for 3 months Probiotic supplementation led to a decrease in the levels of Prevotella (which is enriched in MCI subjects prior to treatment) and this decrease was associated with an improved cognitive score 182 FMT; 36-week clinical trial including 34 PSP-RS patients (receiving healthy donor FMT) and 34 placebo controls (receiving saline) The group receiving FMT had significantly improved symptoms of depression and anxiety, in addition to increases in levels of butyrateproducing bacteria (Faecalibacterium) post-FMT NU-AGE diet randomized trial, 1279 older adults were included and divided into control or intervention groups, adhering to the diet for 1-year Participants with higher adherence to the NU-AGE diet (Mediterraneanlike diet) showed significant improvements in cognition and episodic memory compared to those with lower diet adherence 184 Experimental Dietary intervention; 2-month administration of either soluble fiber diet or control diet in a mouse model of AD AD mice receiving the soluble fiber diet displayed lower memory impairments and anxiety than WT mice receiving the control diet.Additionally, intestinal morphological alterations were reduced in AD mice receiving the fiber diet, an effect accompanied by restoration of butyrate and propionate production in the gut content 185 FMT; 4-weeks of FMT administration from WT control mice to an aged mouse model of AD FMT from age-matched WT control mice alleviated cognitive deficits and reduced the deposition of amyloid-beta in AD mice; FMT also reversed the alterations in specific microbiota changes seen in the AD mouse model prior to FMT 155 FMT; 2-weeks of FMT from control donor mice to a rotenone-induced model of PD mice PD mice receiving FMT from control donors displayed alleviated motor symptoms and better gastrointestinal function, and restores bloodbrain-barrier impairment 187 Probiotic treatment; oral administration of Bifidobacterium breve MCC1274 in WT mice Thea administration of the B. breve MCC1274 probiotic decreased Adlike pathologies in WT mice; soluble hippocampal amyloid-beta levels and neuroinflammation were decreased in mice receiving the treatment 183Dietary intervention;